The role of DNA methylation in neoplasia is well-established, although its importance is only beginning to be appreciated. Hypermethylation is now being recognized s a major cause of gene inactivation in sporadic cancers, and is being found to play a more important role than mutations, in some cancers. The role of DNA methylation in endometrial carcinoma has not been investigated. Endometrial carcinoma hs the fifth leading cause of cancer in women worldwide, with approximately 150,000 cases diagnosed each year. We hypothesize that DNA methylation plays an important role in the pathogenesis of endometrial carcinoma, and propose to study four cancer-related genes, hMLH1, HMSH2, PTEN, and P16 for hypermethylation. We have available for our study a very well characterized population-based series of 316 endometrial carcinoma patients, collected over a period of 10 years, and for whom follow-up data is available. Methylation analysis of the mismatch repair genes, hMLH1 and hMSH2, will be tested, particularly in tumors that demonstrate microsatellite instability. Twenty percent of sporadic endometrial carcinoma tumors demonstrate microsatellite instability, but only a very small proportion of these are due to mutations in the mismatch repair genes. We hypothesize that hypermethylation of these genes are responsible for their inactivation and the subsequent microsatellite instability. We also propose to analyze the methylation status of two tumor suppressor genes, PTEN and p16, in our population of endometrial carcinoma patients, to determine their role in this disease. The PTEN and p16 genes are both good candidates for hypermethylation analysis in endometrial carcinoma, as mutations in the PTEN gene have recently implicated it in this disease, and inactivation of the p16 gene by hypermethylation appears to be a common finding in many other cancers. Our methylation studies will be complemented by mutation studies to determine the comparative role of mutation and methylation in gene inactivation. We will also study the effect of hypermethylation of our four candidate genes on prognosis. These studies should elucidate the role that DNA methylation plays in endometrial carcinoma, further our understanding of the molecular pathogenesis of this disorder, and establish markers that can be used for its diagnosis and prognosis.